Net-Winged Midge Genus Blepharicera Macquart (Diptera: Blephariceridae) in China: The First DNA Barcode Database with Descriptions of Four New Species and Notes on Distribution

SIMPLE SUMMARY: Blephariceridae is a small but highly specialized family in Diptera with extreme morphological adaptations for living in fast-moving waters. Among the seven blepharicerid genera in China, Blepharicera Macquart, 1843 is the species richest genus with 12 known species. There is a serious gap in knowledge concerning larval stages and connecting them to adult morphology. At present, DNA barcoding using mitochondrial (mt) cytochrome c oxidase subunit 1 gene (COI) has proven to be successful for identification in many animal groups. There are about 60 species of the genus Blepharicera known in the world. However, only one identified Blepharicera species from Europe is currently available in DNA sequence repositories, and a DNA barcode database is still lacking for Chinese Blepharicera. To fill this gap, we started a project with the objectives to collect specimens of Blepharicera across China, identify the species by morphological characters, and build an mt COI barcode database for this important insect group. Molecular and morphological markers show that four morphospecies examined represent members of the genus Blepharicera and constitute four new, distinct species. This initial attempt indicates the validity of mt COI barcodes for Blepharicera in delimiting species and contributes to the growing library of DNA barcodes of net-winged midges of the world. ABSTRACT: Mitochondrial (mt) cytochrome c oxidase 1 (COI) gene is more and more widely used for DNA barcoding, which provides a rapid and timely identification as this technique is not limited by polymorphism, sex, and life stages and fundamentally complements traditional evolutionary taxonomy. The present study generated 33 mt COI sequences of seven Chinese Blepharicera Macquart, 1843 species with an average of 594 bp, which represent the first DNA barcode database for Chinese Blepharicera. Genetic distance analysis reveals that intraspecific distances in the genus are generally less than 1.7%, and interspecific distances range from 5.4% to 20.3%. Phylogenetic analysis shows that each species recovered in our analyses is separated from all neighboring species. Based on molecular and morphological data, four Blepharicera species from China, B. beishanica sp. nov., B. dushanzica sp. nov., B. nigra sp. nov. and B. xinjiangica sp. nov., are described and illustrated as new to science. Identification keys for adults and larvae of Chinese Blepharicera are also presented. Geographical analysis shows that Southwest China is the species’ richest region. Our results will be useful in tackling taxonomic problems, understanding species distribution, and resolving nomenclature conflicts associated with Blepharicera species.